Comparative gas phase and pulsed liquid phase sequencing on a modified Applied Biosystems 477A sequencer.
A simple and inexpensive modification of the Applied Biosystems 477A sequencer, to run in the pulsed liquid-phase and in the gas-phase mode of the Edman chemistry, is described. This modification is especially useful for sequencing samples on polyvinylidene difluoride (PVDF) membranes. Additional carriers are required if a sample on a PVDF membrane is sequenced with the pulsed liquid-phase degradation program of the 477A. In the gas-phase mode no such carriers are needed. This eliminates time-consuming preconditioning sequencer cycles and reduces the sequencer background. In addition, initial coupling yields in the gas-phase mode exceeded those in the pulsed liquid-phase mode, whereas the average repetitive yields were similar. Samples spotted onto glass fiber filters pretreated with polybrene and samples spotted or electroblotted onto PVDF membranes were examined. A number of advantages of the gas-phase mode are presented.
Baumann
M
M
University of Helsinki, Department of Medical Chemistry, Finland.
D000595:Amino Acid Sequence; D004591:Electrophoresis, Polyacrylamide Gel; D005740:Gases; D005898:Glass; D006583:Hexadimethrine Bromide; D007782:Lactoglobulins / Q000737:chemistry; D008567:Membranes, Artificial; D008969:Molecular Sequence Data; D009113:Muramidase / Q000737:chemistry; D011145:Polyvinyls; D011506:Proteins / Q000737:chemistry*; D014269:Trifluoroacetic Acid; D005465:Fluorocarbon Polymers
D005740:Gases; D006583:Hexadimethrine Bromide; D007782:Lactoglobulins; D008567:Membranes, Artificial; D009113:Muramidase; D011145:Polyvinyls; D011506:Proteins; D014269:Trifluoroacetic Acid; C009753:fiberglass; C024865:polyvinylidene fluoride; D005465:Fluorocarbon Polymers